Molecular analysis and characterization of nine novel CTSK mutations in twelve patients affected by pycnodysostosis

Abstract

Molecular characterization of twelve unrelated patients affected by the autosomal recessive osteosclerotic skeletal dysplasia, Pycnodysostosis (cathepsin k deficiency), revealed 11 different genotypes. The mutational profile consisted of 12 different mutations, including nine previously unreported ones, spread throughout the whole gene. One mutation occurred in regions coding predomain, two affected the prodomain and nine others occurred in the mature domain. The novel lesions consisted in six missense mutations c.20T>C (p.L7P), c.494A>G (p.Q165R), c.580G>A (p.G194S), c.746T>C (p.I249T), c.749A>G (p.D250G), c.955G>T (p.G319C), two frameshifts c.60_61dupGA (p.I21RfsX29), c.282dupA (p.S95VfsX9) and a splicing mutation c.890G>A (r.785_890del). The six new missense mutations were examined by western blots of COS-7 cells transfected with mutant CTSK genes. The L7P, occurring within the predicted hydrophobic domain of signal peptide, showed a significantly reduced expression level compared to the wild type control. These findings suggested that the mutation affected targeting and translocation of the nascent lysosomal protein across the endoplasmatic reticulum membrane. The novel amino acid changes were also modeled into the three-dimensional structure that predicted incorrect protein folding for all of them. Molecular characterization of the patients is of particular value for genetic counseling of patients and their families as diagnosis of Pycnodysostosis based on enzyme assay is unpractical and thus not offered routinely.

Current research on pycnodysostosis

“…The disorder is caused by a homozygous or compound heterozygous mutation in cathepsin K (CTSK), which is a lysosomal cysteine protease that is highly expressed in osteoclasts. CTSK is involved in the degradation of bone matrix proteins, type I and type II collagen, osteopontin, and osteonectin at a low pH (1)(2)(3)(4)(5)(6)(7). To date, forty-five different CTSK mutations have been reported, including nonsense, missense, frameshift, and splice site mutations as well as small deletions, small and big insertions (Alu sequence) ( Figure 1).…”

“…The condition is also known as Maroteaux-Lamy syndrome and is characterized by a short stature, acroosteolysis of the distal phalanges, dysplasia of the clavicle, osteosclerosis with increased bone fragility, and delayed closure of sutures (1)(2)(3)(4)(5). French artist Henri de Toulouse Lautrec (1864-1901) was suggested to have this condition since he exhibited several phenotypic features of the disorder such as a short stature, parental consanguinity, facial dysmorphism, frequent bone fractures, and large fontanels (8).…”

The Clinical Rehabilitation and Cathepsin K Gene Analysis of a Turkish Family With Pycnodysostosis.

“…T>C transition at exon 2 leads the change of the amino acid leucine (l) to amino acid proline (p). This amino acid change is considered to affect the signal peptide sequence of protein, as indicated in other studies (15).…”

Osteomyelitis in Pycnodysostosis – Report of 2 Clinical Cases

“…This anomaly consists of 12 different mutations [4,5] that produce mutational changes in a lysosomal cystine protease, cathepsin K, the expression of which is reduced in the osteoclasts of these patients [4][5][6][7]. This protease is responsible for degrading collagen type 1, that constitutes 95% of the organic bone matrix.…”